Bladder cancer is the second most common urological malignancy in the United States, which accounts for approximately 13,000 deaths and new cases per year. Ninety percent of bladder malignancies are classified as transitional cell carcinomas (TCCs), which originate from the transitional urothelium that is composed of 3-6 layers thick of basal, intermediate and multinucleate umbrella cells. Seventy percent of tumors are low grade non-invasive papillary lesions at diagnosis, which commonly recur, only 15-20% of which progress into muscle invasive disease. On the other hand, ˜30% of TCCs are muscle invasive at diagnosis. Fifty percent of patients with invasive TCCs die from metastasis within 2 years, and the 5-year survival-rate for metastatic bladder cancer is only 6%.
It has long been observed that patient bladder tumors contain intratumoral heterogeneity, consisting of tumor cells with diverse histological morphologies and distinct biological properties. Primary bladder tumor cells possess differential ability for anchorage independent growth, a property defining the transforming ability of cancer cells. This is supported by evidence that only ˜0.7% of primary bladder tumor cells have the ability to form colonies in the classical two-layered soft-agar assay, which assays for anchorage independent growth properties of tumor cells. In addition, bladder tumor cells demonstrate diverse proliferation and differentiation status, mostly supported by histological analysis showing distinct staining patterns of proliferation markers (i.e. Ki67 and PCNA) and keratin markers defining differentiation status (i.e. K5/14, K8 and K20).
Basic cancer research has focused on identifying the genetic changes that lead to cancer. This has led to major advances in our understanding of the molecular and biochemical pathways that are involved in tumorigenesis and malignant transformation. However, our understanding of the cellular biology has lagged. A large body of literature has examined the effects of particular mutations on the proliferation and survival of model cells, such as cell lines, fibroblasts and most recently in primary epithelial cells; however, the target cell accumulating actual mutations remained to be elucidated.
A tumor can be viewed as an aberrant organ initiated by a tumorigenic cancer cell that acquired the capacity for indefinite proliferation through accumulated mutations. In this view of a tumor as an abnormal organ, the principles of normal stem cell biology can be applied to better understand how tumors develop. Many observations suggest that analogies between normal stem cells and tumorigenic cells are appropriate. Both normal stem cells and tumorigenic cells have extensive proliferative potential and the ability to give rise to new (normal or abnormal) tissues. Both tumors and normal tissues are composed of heterogeneous combinations of cells, with different phenotypic characteristics and different proliferative potentials.
Because most tumors have a clonal origin, the original tumorigenic cancer cell gives rise to phenotypically diverse progeny, including cancer cells with indefinite proliferative potential, as well as cancer cells with limited or no proliferative potential. This suggests that tumorigenic cancer cells undergo processes that are analogous to the self-renewal and differentiation of normal stem cells. Tumorigenic cells can be thought of as cancer stem cells that undergo an aberrant and poorly regulated process of organogenesis analogous to what normal stem cells do. Although some of the heterogeneity in tumors arises as a result of continuing mutagenesis, it is likely that heterogeneity also arises through the aberrant differentiation of cancer cells.
It is well documented that many types of tumors contain cancer cells with heterogeneous phenotypes, reflecting aspects of the differentiation that normally occurs in the tissues from which the tumors arise. The variable expression of normal differentiation markers by cancer cells in a tumor suggests that some of the heterogeneity in tumors arises as a result of the anomalous differentiation of tumor cells. Examples of this include the variable expression of myeloid markers in chronic myeloid leukaemia, the variable expression of neuronal markers within peripheral neurectodermal tumors, the variable expression of milk proteins or the estrogen receptor within breast cancer, and the differential expression of cytokeratins in a wide spectrum of epithelial tumors including bladder cancer.
It was first extensively documented in acute myeloid leukaemia that only a small subset of cancer cells is responsible for the tumor-initiating potential and maintain the ability to self-renew. Because the differences in clonogenicity among the leukemia cells mirrored the differences in clonogenicity among normal hematopoietic cells, the clonogenic leukemic cells were described as leukemic stem cells. It has also been shown for solid cancers that the cells are phenotypically heterogeneous and that only a small proportion of cells are tumorigenic and can self-renew in vivo. Just as in the context of leukemic stem cells, these observations led to the hypothesis that only rare cancer stem cells exist in epithelial tumors.
In support of this hypothesis, recent studies have shown that, similar to leukemia and other hematologic malignancies, tumorigenic and non-tumorigenic populations of breast cancer cells can be isolated based on their expression of cell surface markers. In many cases of breast cancer, only a small subpopulation of cells had the ability to form new tumors. This work strongly supports the existence of CSC in breast cancer. Further evidence for the existence of cancer stem cells occurring in solid tumors has been found in central nervous system (CNS) malignancies. Using culture techniques similar to those used to culture normal neuronal stem cells it has been shown that neuronal CNS malignancies contain a small population of cancer cells that are clonogenic in vitro and initiate tumors in vivo, while the remaining cells in the tumor do not have these properties.
Stem cells are defined as cells that have the ability to perpetuate themselves through self-renewal and to generate mature cells of a particular tissue through differentiation. In most tissues, stem cells are rare. As a result, stem cells must be identified prospectively and purified carefully in order to study their properties. Perhaps the most important and useful property of stem cells is that of self-renewal. Through this property, striking parallels can be found between stem cells and cancer cells: tumors may often originate from the transformation of normal stem cells, similar signaling pathways may regulate self-renewal in stem cells and cancer cells, and cancers may comprise rare cells with indefinite potential for self-renewal that drive tumorigenesis.
The presence of cancer stem cells has profound implications for cancer therapy. At present, all of the phenotypically diverse cancer cells in a tumor are treated as though they have unlimited proliferative potential and can acquire the ability to metastasize. For many years, however, it has been recognized that small numbers of disseminated cancer cells can be detected at sites distant from primary tumors in patients that never manifest metastatic disease. One possibility is that immune surveillance is highly effective at killing disseminated cancer cells before they can form a detectable tumor. Another possibility is that most cancer cells lack the ability to form a new tumor such, that only the dissemination of rare cancer stem cells can lead to metastatic disease. If so, the goal of therapy must be to identify and kill this cancer stem cell population.
The prospective identification and isolation of cancer stem cells will allow more efficient identification of diagnostic markers and therapeutic targets expressed by the stem cells. Existing therapies have been developed largely against the bulk population of tumor cells, because the therapies are identified by their ability to shrink the tumor mass. However, because most cells within a cancer have limited proliferative potential, an ability to shrink a tumor mainly reflects an ability to kill these cells. Therapies that are more specifically directed against cancer stem cells may result in more durable responses and cures of metastatic tumors.
Epithelial tumors contain a mixed population of cancer cells. It has been hypothesized that functional heterogeneity rather than cellular heterogeneity may account for the fact that not all of the cancer cells in solid tumors have a similar ability to drive tumor formation. Mortality from these diseases remains high due to the development of distant metastasis and the emergence of therapy resistant local and regional recurrences. It is therefore essential that we develop a deeper understanding of the biology of this disease in order to develop more effective therapies.
Cancer stem cells are discussed in, for example, Pardal et al. (2003) Nat Rev Cancer 3, 895-902; Reya et al. (2001) Nature 414, 105-11; Bonnet & Dick (1997) Nat Med 3, 730-7; Al-Hajj et al. (2003) Proc Natl Acad Sci USA 100, 3983-8; Dontu et al. (2004) Breast Cancer Res 6, R605-15; Singh et al. (2004) Nature 432, 396-401.